We report here the identification and characterization of mouse mTERF encoding a mitochondrial transcription termination factor. A full-length mTERF cDNA has been isolated and the genomic organization of mTERF has been elucidated. The mouse mTERF gene containing two exons encodes a 380 residue protein with a strong homology to the mTERF-like proteins of human and other organisms, related to mitochondrial transcription termination. Northern blot analysis detected both 1.4 and 5.4kb transcripts. The mouse mTERF 1.4kb transcript agreeing with the size of cDNA is predominately expressed in heart and liver, but at extremely low level in other tissues. In addition, a approximately 5.4kb transcript likely resulting from the retention of intron appears to express abundantly in heart and skeletal muscle, but at very low level in other tissues. Furthermore, immunofluorescence analysis of NIH3T3 cells expressing mTERF-GFP fusion protein demonstrated that the mouse mTERF localizes in mitochondrion. These observations suggest that the mouse mTERF is an evolutionarily conserved mitochondrial transcription termination factor, thereby promoting the termination of transcription in mitochondrial RNA.